Cytosolic calcium regulation by ATP receptors on Ehrlich ascites tumor cells and cardiac ventricular myocytes
Abstract
ATP receptors on Ehrlich ascites tumor cells and cardiac ventricular myocytes were characterized according to their agonist specificities and effects on cellular Ca$\sp{2+}$ homeostasis and found to differ significantly. Addition of 0.4-25 $\mu$M ATP to a suspension of Ehrlich ascites tumor cells loaded with the fluorescent Ca$\sp{2+}$ indicator quin2 or fura2 increased cellular cytosolic (Ca$\sp{2+}$) from 75 nM to 1 $\mu$M. No response was stimulated by adenosine, AMP or ADP, and ATP $>$ ITP, UTP $>$ GTP. The non-hydrolyzable ATP analogue 5-adenylimidodiphosphate was less potent than ATP. ATP did not permeabilize the cells, but released Ca$\sp{2+}$ from intracellular stores and increased Ca$\sp{2+}$ influx. A cardiac ventricular cell preparation was subsequently developed for studying ATP receptor activation in heart. The myocytes maintained a high intracellular ATP:ADP ratio, and responded to cytosolic Ca$\sp{2+}$ effectors including ryanodine, veratridine, norepinephrine, KCl and micromolar concentrations of ATP. (Ca${\sp{2+}}$) transients were not stimulated by adenosine, AMP, or ADP. No nucleotide triphosphate was effective. 2-methylthio ATP or ATP$\gamma$S stimulated a response, partly consistent with a P2y receptor classification. Adenosine tetraphosphate was an antagonist, and the (Ca$\sp{2+}$) response to ATP required Mg$\sp{2+}$. The cardiac ATP receptor was shown to affect cellular Ca$\sp{2+}$ homeostasis through three mechanisms: (1) activation of the L-type voltage-sensitive Ca$\sp{2+}$ channel, (2) release of Ca$\sp{2+}$ from the sarcoplasmic reticulum (SR), and (3) Ca$\sp{2+}$ influx through the Na$\sp{+}$/Ca$\sp{2+}$ exchanger. Extracellular Ca$\sp{2+}$ was required for release of Ca$\sp{2+}$ from the SR. Ca$\sp{2+}$ influx through the Na$\sp{+}$/Ca$\sp{2+}$ exchanger was phosphate dependent and may result from ATP stimulation of Na$\sp{+}$/phosphate cotransport. Cytosolic (Ca${\sp{2+}}$) responses to ATP in myocytes were greatly increased by pretreatment of the cells with norepinephrine, which is costored with ATP in sympathetic neurons. This potentiation is mediated by the $\beta$-adrenergic receptor and does not require intracellular Ca$\sp{2+}$ stores. The effects of ATP on cardiac cellular Ca$\sp{2+}$ homeostasis may significantly alter cell function following exposure to ATP from neural stimulation or nearby cell damage.
Subject Area
Pharmacology|Physiology
Recommended Citation
De Young, Mary Beth, "Cytosolic calcium regulation by ATP receptors on Ehrlich ascites tumor cells and cardiac ventricular myocytes" (1990). Dissertations available from ProQuest. AAI9026545.
https://repository.upenn.edu/dissertations/AAI9026545